A polymerase chain reaction (PCR) is a method of amplifying DNA by multiple synthesis of a selected region of the DNA, thereby producing a large amount of DNA by cloning a very small amount of the DNA.
Through the PCR, only a desired segment of DNA may be amplified from very large DNA like genomic DNA. The PCT generally includes denaturation, primer annealing and DNA polymerization processes.
Recently, real-time PCR is well known to those of ordinary skill in the art. The real-time PCR is a technology which allows monitoring of a reaction state in real-time by measuring an intensity of fluorescence showing the level of DNA amplification at every cycle in a status that a reaction product in a gel is not separated by electrophoresis. Therefore, in the real-time PCR, there are some advantages in that precise quantitative analysis is allowed, and it is possible to simply and rapidly perform the analysis without the electrophoresis, and also there is less risk of contamination.
A real-time PCR apparatus includes a thermal cycler for PCR and a fluorometer for detecting fluorescence of a reaction product. A conventional real-time PCR apparatus is comprised of a thermoelectric element, a thermal block for transferring heat to a reaction tube in which a sample is received, a light source for irradiating excitation light to the sample in the tube, and a light receiving part for receiving the fluorescence generated from the sample. In the conventional real-time PCR apparatus, cooling and heating cycles are repeatedly performed by using the thermoelectric element so as to react the sample, and the excitation light is irradiated to the sample using the light source and the light receiving part at every end of each cycle, and then an amount of the fluorescence generated from the sample is measured so as to display the progress of the PCR in real-time.
However, in the conventional real-time PCR apparatus, it is possible to treat a plurality of samples, but it is impossible to successively react the samples at regular time intervals, and also it is impossible to provide other samples in the reaction tube during the reaction of the sample until the reaction is completed.
To solve the above problems, there has been proposed various continuous real-time monitoring apparatuses.
In U.S. Pat. No. 6,033,880, there is disclosed a PCR apparatus using a capillary tube. According to the PCR apparatus, a heat transfer block includes four constant temperature blocks, and samples and reagents are supplied to or removed from the capillary tube using a solution supplying unit. The PCR is performed by rotating the heat transfer block and changing temperature transferred to the capillary tube using the above-mentioned apparatus. The problem in this type apparatus is that the heat transfer block should be rotated to perform the PCR, and also the reproducibility of the PCR is deteriorated since the PCR may be changed depend on a contacting level between the capillary tube and the heat transfer block.
Further, in this type apparatus, it is impossible to perform the PCR at time intervals. Furthermore, since the above apparatus can measure the progress of the reaction only after completion of the PCR, there is another problem that a user cannot check the progress of the reaction before the completion of the PCR.
To solve the above problems, there has been proposed a new PCR real-time monitoring apparatus in Korean Patent No. 593263 (titled “a high throughput device for performing continuous-flow reactions”), in which a temperature circulating unit for PCR, comprised of a capillary tube and a circular heating block, is provided.
In this apparatus, the capillary tube of 3.5 meters in length is wound 33 times on a copper block of 30 mm in diameter, which is divided into melting, annealing and extension temperature regions. When a reaction mixture flowed in the capillary tube is circulated once around the heating block formed of copper, each cycle of the PCR is performed. In this method, the capillary tube through which the PCR sample is flowed is wound on the heating block, and the capillary tube is scanned by a scanning unit having a fluorescence detector. Thus, the scanning unit is a means for irradiating light to the capillary tube wound on the heating block using a light irradiating unit and measuring an amount of fluorescence generated in the capillary tube.
According to the above-mentioned method, the light irradiating unit for irradiating light to the capillary tube wound on the heating block and a sensor for measuring the fluorescence generated from the capillary tube are installed at a moving stage, so that the scanning unit is linearly driven and the light is irradiated, in turn, to the capillary tube according to movement of the scanning unit. Then, the fluorescence generated from the capillary tube is measured, in turn, according to the movement of the scanning unit.
In the above-mentioned technology, there is disclosed the scanning unit in which a fluorescence detecting sensor and a light source for generating a light beam having a desired wavelength are moved at a constant speed above the heating block on which the capillary tube is wound. The light source and the fluorescence detecting sensor installed at the scanning are moved in an axial direction that is parallel with a central axis of the heating block on which the capillary is wound or that is cross the central axis thereof, so as to irradiate the light to the capillary tube or measure the fluorescence. Whenever perform the scanning, monitoring of the PCR is performed once, and the multiple capillary tubes are scanned upon the scanning. In order to scan the fluorescence generated from the sample in the capillary tube while the light source and the fluorescence detecting sensor installed at the moving stage are moved at a constant speed, it is necessary to provide a motor, a conveying unit like liner conveying means, a conveying guide unit, driving means for providing power the conveying unit and so on. However, since the light source including a plurality of optical lenses uses an expensive lens like an object lens and it is also necessary to precisely arrange the light source and the fluorescence detecting sensor in order to precisely control an optical path, there is a problem that a manufacturing cost of the PCR apparatus is remarkably increased. In addition, since the PCR apparatus includes the plurality of lenses, the conveying unit, the power transferring unit, the driving means and the like, this may cause increase of its size and malfunction thereof.
Therefore, there is necessity of providing a new continuous PCR real-time monitoring apparatus which solves the above-mentioned problems and has an excellent and economical real-time monitoring effect.